Porous cellulose beads provide a relatively lowcost, stable material possessing versatile chemical properties such that thay can be useful as a carrier for immobilized enzymes and other active biological agents.
While ordinary cellulose particles and regenerated cellulose powders meet most of the desired requirements of good carriers to which enzymes can be immobilized, they suffer from configural disadvantages which cause column reactors to become tightly packed resulting in reduction of flow and sometimes channeling, and thus insufficient contact between the immobilized enzyme and reaction fluid. The immobilization of enzymes on an insoluble carrier is a widely-accepted technique for a practical application of enzymes, avoiding the necessity of employing fresh enzymes for each desired use. Through immobilization of the enzyme, stabilization is achieved which provides for efficient enzyme use and provides for the design and operation of enzyme reactors in a continuous mode.
To a large degree, the success of an immoblized enzyme for use in practical application depends upon the properties of the carriers employed for immoblization. Accordingly, a good carrier should meet the requirements of being inexpensive and should be of such a physical shape that it is easy to be employed in reactors. In this regard, the shape of a spherical bead is particularly desirable, since it is useful in a packed bed, fluidized bed, expanded bed, stirred tank, or other common types of chemical reactor designs. Such a carrier should also have the proper physical and mechanical strength such that it will not be crushed or deformed when packed in a tall column. Crushing and deformation results in the column becoming tightly packed, thereby blocking the flow of liquid reagents through the column, thus decreasing the efficiency of the chemical reactor. Suitable carriers should also possess versatile chemical properties such that the immoblization of enzymes and other biological agents onto the carrier through ionic or chemical covalent bonding, as well as surface absorption, can be readily achieved. In this regard, the carrier should have a high capacity for forming a large number of bonds such that each unit of the carrier can immobilize large amounts of the enzyme desired. Thus, a carrier having a high degree of porosity and uniformly distributed internal void spaces is particularly desirable. Such porosity provides for good diffusion of chemical reagents or reaction products into and out of the internal void spaces of the cellulose beads. Carriers should be chemically stable, physically strong, and made of inert material which resists microbiological attack causing carrier deterioration in order to provide an immobilized enzyme system having a prolonged active life.
Currently, porous glass and porous ceramic particles are commonly employed for the immoblization of enzymes and while such particles meet most of the above requirements for an acceptable particle, they are relatively expensive. Furthermore, the number of chemical reactions which may be used for immobilization of enzymes to glass and ceramic carriers is limited.
In U.S. Pat. Nos. 3,947,325; 3,905,954; 3,573,277; 3,505,299; 3,501,419; 3,397,198; 3,296,000; 3,251,824; 3,236,669; 2,843,583; 2,773,027; 2,543,928 and 2,465,343, there is described the preparation of a variety of cellulose materials in a variety of forms, some of which are described as suitable for use in fixing biologically-active materials such as enzymes or ion-exchange groups thereto. However, these processes seem to suffer also from the disadvantage of being expensive and the products obtained generally are of an undesirable physical shape for use in such chemical reactors as packed beds and fluidized beds. In particular, the prior art fails to provide a means for producing spherical shaped cellulose beads having a uniform distribution of pores throughout the surface and a large uniformly porous internal void space. Furthermore, the cellulose particles and powders of the prior art generally are of such a small particle size that they are not suited for use in chemical reactors. In addition, the cellulose powders and particles of the prior art often have a hard surface skin which causes severe diffusional hinderance and inefficient use in chemical reactors.
In our earlier application, we describe the process of making highly porous cellulose beads of uniform porosity which were found highly suitable for immobilizing enzymes. We have found that these beads may also be useful in the purification and separation of enzymes, proteins, nucleic acids and the like. Furthermore, the beads may be useful to separate metallic ions from dilute solutions containing same.
Accordingly, the primary object of the present invention is to provide a means for preparing inexpensive, highly-porous, stable particles having versatile chemical properties whereby they may be useful as a carrier to which enzymes or other biologically-active materials can be immoblized.
A further object of the present invention is to provide a method for the transformation of cellulose derivatives into highly-porous particles having good mechanical stability such that it will provide for adequate passage of liquid therethrough when operated in packed bed reactors.
Still yet another object of the present invention is to provide a porous cellulose bead having sufficiently large surface area to provide high immobilization capacity of enzymes.
Still a further object of the present invention is to provide a porous cellulose bead having improved physical and mechanical strength so that it will not be crushed and deformed when used in chemical reactors.
Yet a further object of the invention is to provide an improved means for the purification and/or separation of enzymes, proteins, nucleic acids and the like.
Yet another object of our invention is to provide a means for the separation of metallic ions from dilute solutions containing same.
These and other objects of the present invention will be more fully apparent from the discussion set forth hereinbelow.